Mammalian Cell Expression System

High expression level, easy operation, short circle, low cost and easy cultivation with high density and large-scale are the advantages of prokaryotic expression system. To complete antibody and glycoprotein biological drugs, the synthesis, secretion ,bio-activation, stability in vivo and immunogenicity of the expression product are often affected by the folding of the peptides , the forming of disulfide bond , the type of glycosylation and whether has glycosylation or not. The product is non- glycosylated and usually exist in form of inclusion body , because the prokaryotic cell has no endoplasmic reticulum and Golgi apparatus which are related to glycosylation. Although the yeast , insect and other eukaryotic cell has the modification of glycosylation, the way of glycosylation is different from that of the human and the other mammalian cell does. Compare with other eukaryotic cell expression system, the proteins from target gene expressed in mammalian cell are nearly the same with the nature proteins in the term of structure, glycosylation style and the way of glycosylation, and can be assembled into multi-subunit protein. Moreover, the mammalian cell can be cultivated in suspension way or in serum-free medium with high density which the volume could reach to 1000 L and even more .

In conclusion , the advantages of mammalian cell expression system are forming right folding of protein, having many post-translation processing functions such as carrying out complex N－style glycosylation , accurate O –style glycosylation and so on. Therefore, product from mammalian cell is the most close to the nature proteins of advanced creature in the respect of the molecular structure, character of physics and chemistry , and biological function.

Service content

Character and advantage

Expression vector construction

Cell transient transfection（Calcium phosphate coprecipitation,lipofectin transfection, electroporation）：short circle，protein with high activity. We can supply experiment data and the purified product.
Stable transfection cell line（antibiotic screen）：the target gene could do sustained expression with high protein activity . We can supply experiment data and the purified product.Continue to order the same protein can enjoy the 30% ~50% discount.

Expression condition optimization

Protein expression and purification

Transient transfection of mammalian cell

Experiment content

Experiment flow

Content of supply

Time

1.Prokaryotic clone

Obtain the gene

Nucleic Acid Electrophoresis picture

2 weeks

PCR product connect to the redesigned vector pcDNA3.1or psv2-dhfr

Transformation into DH5α

Large-scale extract the recombinant plasmid

2.Transient transfection

Cells are moved into a plates and cultured
After the total cells reach in 70%~80% of the hole ，start to do transfection
48~72h later harvest the cells

The experiment data of Transfection

3~4 days

3.Protein expression and purify

By western blot test

Western blot

4 weeks

Purify the expression product

The purified product of 1L supernatant

Stable transfection of mammalian cell

Service content

Experiment flow

Content of supply

Time

1. Prokaryotic clone

Obtain the gene

Nucleic Acid Electrophoresis picture

2 weeks

PCR product connect to the redesigned vector pcDNA3.1or psv2-dhfr

Transformation into DH5α

Large-scale extract the recombinant plasmid

2 Stable transfection

Do antibiotic screening after transfection，construct the stable transfection cell line

The experiment data of transfection and the cell strain with sustained and stable expression

3~5 months

3. Protein expression and purify

By western blot test

Western blot

2 weeks

Purify the expression product

The purified product of 1L supernatant

We have special vectors which have the advantages of quick cloning target gene , facilitating purification and improving the expression level of the interest protein.